Hydrogels are a class of materials that have significant promise for use in soft tissue and bone engineering. The general characteristic of hydrogels that make them important materials for these applications are their well hydrated, porous structure. The present invention provides a new class of environmentally responsive peptide-based hydrogels that fulfill critical material requirements not currently met with existing technology. Hydrogels of the invention may be designed to be compatible with the adhesion and proliferation of various cell types, e.g., fibroblasts and osteoblasts, making them potential tissue engineering scaffolds for generating connective tissue and bone. There is a demanding set of biological and material properties required of hydrogels for use in tissue regeneration. Irrespective of ultimate target tissue type, a hydrogel must exhibit a general set of biological properties. First, the material must be cytocompatible. Cytocompatibility, defined herein, means that the hydrogel must not be cytotoxic to desired cells. Second, the material must be biocompatible. Biocompatible, defined herein, means that a scaffold does not cause a significant immunological and inflammatory response if placed in vivo for tissue regeneration and is preferably biodegradable affording non-toxic species. The present invention relates to the development of new materials using novel self-assembly methodology and the assessment of resultant material cytocompatibility.
Desired material properties are challenging to comprehensively incorporate into any one material since some desired properties are seemingly mutually exclusive. For example, the morphology of an ideal hydrogel contains a high level of porosity (spanning nanoscale to microscale dimensions) for cell motility and nutrient/waste diffusion. Also, the hydrogel should primarily be composed of aqueous media with as little solid material as possible in order to allow ample volume for cell proliferation and ease of eventual scaffold biodegradation. However, despite their dilute, porous nature, these well hydrated materials must also be mechanically rigid. This apparent contradiction, rigidity from a dilute porous scaffold, must be inherently addressed by the design of constituent molecular crosslinks (chemical and/or physical) formed during the hydrogelation process. However, introducing chemical crosslinks may be biologically problematic since by-products from the crosslinking chemistry may be toxic and difficult to remove from the scaffold. Ideally, benign, biocompatible chemical or physical crosslinking methods should be used for either in vitro gelation for eventual incorporation in the body or direct, rapid in vivo gelation where formation of crosslinks are triggered by physiological stimuli (temperature, ionic strength, pH, etc). The idea of using environmental triggers to initiate material formation via self-assembly is being actively pursued. For example, it has been shown that peptide self-assembly (and thus gelation) can be triggered by the release of salt from temperature and light sensitive liposomes (Collier, et al., Journal of the American Chemical Society 2001, 123, 9463-9464). An additional design complication is that hydrogel rigidity seemingly precludes any viable processability in preformed scaffolds. For example, one may wish to form a rigid tissue engineering construct in vitro but subsequently inject it into a host for tissue regeneration. Injection is not possible in a permanently crosslinked, rigid network.
Current hydrogel technology utilizes both naturally-derived macromolecules and synthetic polymers. Generally, hydrogels prepared from natural polymers exhibit favorable biological properties but may lack desired material properties, e.g. low sample rigidity. In contrast, synthetic polymers can be engineered for desired material properties but may display limited cytocompatibility. A common approach to increase the cytocompatibility of synthetic polymers is to incorporate peptide epitopes, for example RGD motifs. However, incorporating these motifs into preformed polymers in a regiospecifically controlled manner is extremely difficult to impossible. As a result, controlling the material properties of the polymers is problematic. For example, controlling the concentration of an epitope displayed on the surface for cell adhesion or controlling the accessibility of the epitopes is a challenge. In addition, these scaffolds are structurally homogeneous (not porous) on the microscale due to their underlying molecular network structure, which can limit cell proliferation. These systems must undergo additional processing (e.g. freeze-thaw cycling, particulate leaching, microsphere sintering and non-woven fiber formation) in order to introduce microscale porosity in the gel network. In short, there is currently no single hydrogel system that successfully incorporates all requisite properties of an ideal tissue engineering scaffold.
An opportunity and need exists for the design of novel hydrogel scaffold strategies. There remains a need for rigid, porous, easily processed, cytocompatible hydrogels that can be rapidly formed in vitro or in vivo.